The cobalamin family comprises vitamin B12 (cyanocobalamin) and its axial-ligand substituted congeners, such as hydroxocobalamin, methylcobalamin and adenosylcobalamin, among others. Various cobalamins have been used effectively for the treatment of conditions resulting from cobalamin deficiency, such as hematological abnormalities (e.g., macrocytosis and megaloblastic anemia) and neurological impairments (ranging from neuropathy and demyelination to confusional states, mood shifts, memory loss, dementia and depression). These classical sequelae to chronic vitamin B12 deficiency and their treatment are well known (Schneider and Stroinski, Comprehensive B12 (New York: Walter de Gruyter & Co., 1987)). In addition, a number of other diseases or disease states have been characterized by some form of cobalamin deficiency; in many of these cases cobalamin treatment has been reported to result in an amelioration of symptoms or other improvement in the patient's condition. The diseases and disease states studied include anemias of various kinds, autoimmune conditions, disorders of carbohydrate and lipid metabolism such as diabetes and atherosclerosis, neuropathies of various etiologies, mitochondrial disorders and/or deficiencies of cellular bioenergetics, neurodegenerative diseases, mental and psychiatric disorders, endocrine dysfunctions, infertility and reproductive disorders, osteoporosis, immunodeficiencies, AIDS and cancer.
Erythrocyte macrocytosis and macrocytic anemia are often considered to be the classic hematological signs of cobalamin deficiency, especially when found in conjunction with low hemoglobin values. Recently, however, a more complex and varied picture of cobalamin-deficiency anemia has emerged. For example, a surprisingly high rate of incidence of cobalamin deficiency has been detected in sickle cell disease (SCD) patients (Carmel & Johnson, Blood 86, Suppl. 1,644a (1995); Al-Momen, J. Intern. Med. 237, 551-555 (1995)), where the sickle cell anemia may mask a coexisting cobalamin deficiency anemia. The frequent association of folate deficiency with SCD further obscures and complicates the clinical picture. In particular, investigators have concluded that the frequency of cobalamin abnormalities is high enough to warrant concern about the indiscriminate use of folate supplements in SCD (Carmel & Johnson, op. cit.), since folate administration in the absence of cobalamin is known to exacerbate the neuropathology of cobalamin deficiency. Thus, cobalamin supplementation may be especially desirable in those SCD patients who are being treated with folate. Furthermore, an increased unsaturated B12 binding capacity has been unexpectedly found in association with iron deficiency anemia (Rosner & Schreiber, Am. J. Med. Sci. 263, 473-480 (1972)) suggesting an increased need for vitamin B12 under these circumstances. Delayed plasma clearance of radiolabeled cobalamin has also been reported in iron deficiency anemia; one explanation proposed for this effect is a decreased uptake of vitamin B12 by tissues as a result of diminished erythropoiesis (Cook & Valberg, Blood 25, 335-344 (1965)). Since ethrythrocytes appear to play a significant role in delivering cobalamin to tissues (Sorrell et al., Am. J. Clin. Nutr. 24 924-929 (1971)), one may conclude that any cause of anemia resulting in diminished erythropoeisis and/or decreased red cell numbers can induce a state of functional cobalamin deficiency. Therefore, cobalamin supplementation may be useful in treating various forms of anemia and especially in treating those cases associated with coexisting folate deficiency, e.g., as in thalassemia (Kumar et al., Am. J. Clin. Pathol. 84,668-671 (1985)) or SCD.
Pernicious anemia, the prototypical disorder of cobalamin absorption, is generally characterized by gastric atrophy and autoimmune attack on the parietal cells of the gastric fundus, with consequent depletion or impairment of intrinsic factor. Suggestively, an increased prevalence of other autoimmune disorders, such as vitiligo, Graves' disease, Hashimoto's thyroiditis, Type I diabetes, Sjogren's syndrome and rheumatoid arthritis, is found among pernicious anemia patients; the resulting pattern of coexisting autoimmune disease is referred to by the term polyglandular autoimmune syndrome (Leshin, Am. J. Med. Sci. 290, 77-88 (1985)). Many autoimmune disorders, whether components of a polyglandular autoimmune syndrome or not, are associated with abnormal cobalamin metabolism. For example, cases of Sjogren's syndrome, rheumatoid arthritis, systemic lupus erythematosus, dermatomyositis and autoimmune hemolytic anemia have been associated with elevated levels of apo-transcobalamin II, the unbound form of the B12-binding protein which carries cobalamin from serum to cells, thus suggesting an increased demand for vitamin B12 in these conditions (Gimsing et al., Scand. J. Rheumatol. 11, 1-4 (1982); Frater-Schroder et al., Lancet 2, 238-239 (1978)). Furthermore, cobalamin deficiencies have been noted in both synovial fluid (Ono et al., J. Vitaminol. 18, 1-2 (1972)) and serum (Vreugdenhil et al., Ann. Rheum. Dis. 49, 93-98 (1990)) of patients with rheumatoid arthritis, and in the sera of patients with systemic lupus erythematosus (Molad et al., Am. J. Med. 88, 141-144 (1990)), inclusion body myositis and Sjogren's syndrome (Khraishi et al., J. Rheumatol. 19, 306-309 (1992)). A number of these conditions have been shown to be responsive to cobalamin therapy. For example, methylcobalamin has been reported efficacious in the treatment of rheumatoid arthritis (Yamashiki et al., J. Clin. Lab. Immunol. 37, 173-182 (1992)). Likewise, some cases of multiple sclerosis are associated with cobalamin deficiency (Reynolds et al., Arch. Neurol. 48, 808-811 (1991); Baig & Qureshi, Biogenic Amines 11, 479-485 (1995)), and improvement in some patients has been noted upon treatment with cyanocobalamin (Levin, Am. J. Digest. Dis. 22, 96-97 (1955)) or methylcobalamin (Kira et al., Intern. Med. 33, 82-86 (1994)). Both psoriasis and lupus erythematosus have been successfully treated with cyanocobalamin (Stingily, Miss. Doctor 32, 222-223 (1955)), while cases of vitiligo have responded to treatment with vitamin B12 combined with other vitamins (Montes et al., Cutis 50, 39-42 (1992)).
Several distinct lines of evidence connect cobalamin deficiency with diabetes mellitus and other disorders of carbohydrate metabolism. In animals rendered experimentally diabetic, a significant decrease in both serum and tissue cobalamins has been shown to accompany the induction of ketosis (Nath & Nath, J. Vitaminol. 15, 174-177 (1969)). Elevated levels of unsaturated B12 binding capacity (UBBC), a measure of apo-transcobalamins in serum, have been noted in diabetic patients with hyperglycemia, with UBBC normalizing upon restitution of glycemic control (Takahashi et al., Diabetes Res. Clin. Pract. 25, 13-17 (1994)). Moreover, it has long been known that insulin resistance is common among patients presenting with both diabetes and pernicious anemia (Adams, Med. Clin. N. Amer. 8, 1163-1170 (1925); Wright, Clifton Med. Bull. 12, 64-67 (1926)), whereas vitamin B12 administration is effective in increasing insulin sensitivity in some diabetics (Ralli et al., J. Clin. Endocrinol. Metab. 15, 898 (1955)). Likewise, treatment of pernicious anemia patients with cyanocobalamin is known to improve glucose tolerance, an effect attributed to the regulatory influence of vitamin B12 on carbohydrate metabolism (Panzram, Schweiz. Med. Wschr. 91, 234-240 (1961)). Vitamin B12 has also been reported effective in restoring the impaired glucose tolerance induced by thyroid hormone, corticosteroids and by various disease processes (Hadnagy et al., Int. Z. Vitaminforsch. 33, 141-150 (1963)). Since Type II diabetes, obesity, hypertension, coronary artery disease and age-related glucose intolerance have all been associated with increased insulin resistance (Reaven, Diabetes 37, 1595-1607 (1988); Jackson, Diabetes Care 13, Suppl. 2, 9-19 (1990)), cobalamin supplementation is likely to be of specific benefit in these conditions. Alcoholism is another disease associated with impaired glucose tolerance (Dobbins, U.S. Pat. No. 4,918,102 (4/17/90)). Tissue levels of vitamin B12 have been reported depleted in chronic alcoholism (Kanazawa & Herbert, Lab. Invest. 53, 108-110 (1985)), thus offering a rationale for supplementation. In addition, patients with cystic fibrosis (CF) often develop impaired glucose tolerance and progress to frank diabetes characterized by peripheral and hepatic insulin resistance (Hardin et al., Diabetes 44, Suppl. 1, 200A (1995)). CF also is known to be associated with aberrations in vitamin B12 absorption and transport (Lindemans et al., Acta Paediatr. Scand. 74, 795-796 (1985)).
Atherosclerosis involving renal, peripheral and cardiovascular sites is a major complication of diabetes mellitus; hyperinsulinemia and especially hyperglycemia are believed to contribute to the development of atherosclerosis by altering vascular metabolism and by inducing elevated lipid levels (Kunjathoor et al., J. Clin. Invest. 97 1767-1773 (1996)). As noted above, cobalamin supplementation can help normalize insulin sensitivity in diabetic patients and by implication reduce the risk of atherosclerotic involvement. Moreover, vitamin B12 deficiency has been shown to induce hypercholesterolemia in various animals (Hsu & Chow, Fed. Proc. 16, 63 (1957)), whereas treatment with vitamin B12 has been shown to attenuate the increase in serum cholesterol in an animal model of cholesterol-induced atherosclerosis (Nath & Saikia, Arch. Biochem. Biophys. 79, 216-233 (1959)). Obesity is another consequence of impaired glucose tolerance and a frequent concomitant of atherosclerosis. Animals maintained on a cobalamin-deficient diet with normal fat content tend to accumulate fat and become obese, while controls receiving vitamin B12 supplements remain normal (Ling & Chow, in Vitamin B12 und Intrinsic Factor (Stuttgart: Ferdinand Enke Verlag, 1957), 127-132).
Another factor predisposing to atherosclerosis is the accumulation of homocysteine in serum (Malinow, J. Intern. Med. 236, 603-617 (1994)). Cobalamin deficiency results in the failure of the cobalamin-dependent enzyme methionine synthase to remethylate homocysteine to methionine, with a consequent accumulation of homocysteine. Thus, deranged cobalamin metabolism can induce atherosclerotic lesions via elevated serum homocysteine (McCully, Nutr. Rev. 50 7-12 (1992)). Conversely, significant reductions in plasma homocysteine, cholesterol, triglycerides and low density lipoprotein have been observed in patients with ischemic heart disease treated with cyanocobalamin and other nutrients (Olszewski et al., Atherosclerosis 75 1-6 (1989); Olszewski, ibid. 88, 97-98 (1991)). Similarly, elevated plasma homocysteine in diabetic patients has been shown to be associated with clinical macroangiopathy, with reductions in homocysteine levels following upon treatment with methylcobalamin (Araki et al., Atherosclerosis 103, 149-157 (1993)). Cobalamin may be useful in the treatment of other homocysteine-related vascular occlusive disease, such as diabetic retinopathy (Neugebauer et al., Lancet 349, 473-474 (1997)) and arterial and venous thrombosis (Harpel et al., J. Nutr. 126, 1285S-1289S (1996)).
Diabetic neuropathy has been linked with a form of cobalamin deficiency in peripheral nerve (Tanaka et al., in Diabetic Neuropathy (Amsterdam: Excerpta Medica, 1982), 114-119). Effective treatment of diabetic neuropathy has been reported with cyanocobalamin (Sancetta et al., Ann. Intern. Med. 35, 1028-1048 (1951)), methylcobalamin (Yaqub et al., Clin. Neurol. Neurosurg. 94, 105-111 (1992)), and with hydroxocobalamin in combination with other B vitamins (Sakitama et al., J. Nutr. Sci. Vitaminol. 35, 95-99 (1989)). Methylcobalamin has also been found useful in the treatment of autonomic and peripheral neuropathies in uremic patients undergoing hemodialysis (Taniguchi et al., Clin. Ther. 9, 607-614 (1987)). Moreover, treatment with methylcobalamin has been shown to attenuate markedly the incidence of experimental allergic neuritis, an animal model of Guillain-Barre syndrome and other postinfectious and postvaccinal neuropathies (Inada et al., in Vitamin B12 (Berlin: Walter de Gruyter & Co., 1979), 1017-1018). Other peripheral neuropathies which may be associated with cobalamin deficiency and for which cobalamin supplementation has been suggested include leprous neuropathy and the deficiency neuropathy of pellagra (Bedi et al., J. Assoc. Physicians India 21, 473-479 (1973)). In addition, some cases of orthostatic hypotension are known to be due to autonomic neuropathy secondary to cobalamin deficiency (Lossos & Argov, J. Am. Geriatr. Soc. 39, 601-602 (1991)). Likewise, instances of tinnitus (Shemesh et al., Am. J. Otolaryngol. 14, 94-99 (1993)) and optic neuritis (Heaton, Proc. Nutr. Soc. 19, 100-105 (1960)) may represent cases of sensory neuropathy treatable with cobalamin. Remarkably, even neuropathies of a genetic etiology may be treatable with cobalamin. For example, Leber's hereditary optic neuropathy (LHON) is a genetic disease associated with disturbances in cobalamin metabolism (Linnell et al., Clin. Sci. 37, 878 (1969)) and also with defects in mitochondrial DNA and electron transport activity (Rizzo, Neurology 45, 11-16 (1995)). The latter author has proposed that mitochondrial ATP depletion secondary to vitamin B12 deficiency is a metabolic trigger which can precipitate the symptomatology of LHON, and that supplementation with cobalamin (e.g., hydroxocobalamin) can enhance the potential for recovery.
It has long been recognized that cobalamin plays an important role in maintaining mitochondrial integrity (Reddi & Nath, J. Vitaminol. 17, 101-104 (1971)). One explanation for this effect involves the function of the mitochondrial enzyme methylmalonyl-CoA mutase, which utilizes adenosylcobalamin as coenzyme to catalyze the isomeization of L-methylmalonyl-CoA to succinyl-CoA. In cobalamin deficiency the activity of the enzyme is decreased and methylmalonic acid accumulates in plasma and tissues as a result (Toyoshima et al., J. Nutr. 125, 2846-2850 (1995)). Methylmalonic acid is a reversible inhibitor of succinate dehydrogenase, an enzyme which occupies a key locus at the intersection of the tricarboxylic acid cycle and the electron transport chain (Toyoshima et al., op. cit.; Dutra et al., J. Inher. Metab. Dis. 16, 147-153 (1993)). Defects in other components of the electron transport chain have also been noted in cobalamin deficiency (Krahenbuhl et al., J. Biol. Chem. 266, 20998-21003 (1991)), and it has been concluded that cobalamin deficiency can impair ATP synthesis by disrupting cellular bioenergetics (Nakai et al., Pediatr. Res. 30, 5-10 (1991)). This conclusion is significant not only for the understanding it provides of the pathogenesis of LHON (Rizzo, op. cit.), but also because decreased ATP levels and declining mitochondrial membrane potential have been proposed to initiate induction of apoptosis, or programmed cell death (Richter et al., FEBS Lett. 378, 107-110 (1996)). Apoptosis is thought to be involved in the pathogenesis of a number of conditions, including age-related diseases (Wolvetang et al., FEBS Lett. 339, 40-44 (1994)) and retroviral infections such those due to feline immunodeficiency virus (Danave et al., J. Virol. 68, 6745-6750 (1994)) and human immunodeficiency virus (HIV) (Macho et al., Blood 86, 2481-2487 (1995)). Inhibition of succinate dehydrogenase also results in secondary excitotoxicity which may play a role in hypoxia/ischemia and in neurodegenerative disease in general (Davolio & Greenamyre, Neurosci. Lett. 192, 29-32 (1995); Beal, Ann. Neurol. 38, 357-366 (1995)). All of these conditions may be initiated or promoted by cobalamin deficiency and, conversely, ameliorated by cobalamin therapy. Fibromyalgia (Bengtsson et al., Arth. Rheum. 20, 817-821 (1986)), Reye's syndrome (Partin et al., N. Engl. J. Med. 285, 1339-1343 (1971)) and other diseases associated with metabolic, viral, hypoxic or genetic disruption of cellular bioenergetics (Scholte, J. Bioenerg. Biomembr. 20, 161-191 (1988)) may likewise be amenable to treatment with cobalamin.
The neurotropic properties of cobalamins have led investigators to search for cobalamin deficiencies among cases of neurodegenerative disease. Alzheimer's disease has been characterized by low cobalamin levels in serum (Karnaze & Carmel, Arch. Intern. Med. 147, 429-431 (1987)), in cerebrospinal fluid (CSF) (Regland et al., Acta Neurol. Scand. 85, 276-281 (1992)), or in both (Ikeda et al., Acta Psychiatr. Scand. 82, 327-329 (1990)). Treatment with vitamin B12 has resulted in reduction of elevated platelet monoamine oxidase activity (Regland et al., Eur. Arch. Psychiatry Clin. Neurosci. 240, 288-291 (1991)) in Alzheimer's patients. In addition, patients exhibiting high CSF cobalamin levels after treatment with methylcobalamin have shown improvements in intellectual function and memory (Ikeda et al., Clin. Ther. 14, 426-437 (1992)) and in mood and sociability (Mitsuyama, in Basic, Clinical, and Therapeutic Aspects of Alzheimer's and Parkinson's Diseases, Vol. 2 (New York: Plenum Press, 1990), 15-18). Down's syndrome, a condition which eventually manifests a neuropathology resembling Alzheimer's disease in most patients over 40 (Ellis et al., Neurology 24, 101-106 (1974)), is also associated with macrocytosis and low serum B12 (Howell et al., Scand. J. Haemat. 11, 140-147 (1973)). Increases in IQ among children with Down's syndrome have been reported upon supplementation with high daily doses of cobalamin combined with other vitamins (Harrell et al., Proc. Natl. Acad. Sci. USA 78, 574-578 (1981)).
Other neuropathological conditions which may be associated with low serum cobalamin include amyotrophic lateral sclerosis and Parkinson's disease (Bauer & Heinrich, in Vitamin B12 und Intrinsic Factor (Stuttgart: Ferdinand Enke Verlag, 1957), 499-509). Improvement in some cases of amyotrophic lateral sclerosis has been reported upon treatment with vitamin B12 (Levin, Am. J. Digest. Dis. 22, 96-97 (1955); Krolyunitskaya et al., Zhur. Nevropat. Psikhiat. 56, 319-322 (1956)). Cobalamin deficiency has also been linked with excessive production of excitotoxic substances which may play a role in Huntington's disease (Brennan et al., Med. Hypotheses 7, 919-929 (1981)) and other neuropathologies (Santosh-Kumar et al., Med. Hypotheses 43, 239-244 (1994)). Conversely, recent studies have demonstrated a neuroprotective effect of methylcobalamin against the glutamatergic excitotoxicity induced by hypoxia/ischemia or by glutamate itself (Akaike et al., Eur. J. Pharmacol. 241, 1-6 (1993); Yarnamoto et al., Eur. J. Pharmacol. 281, 335-340 (1995)). Glutamatergic excitotoxicity of a related sort has been implicated in the pathogenesis of amyotrophic lateral sclerosis and Alzheimer's, Parkinson's and Huntington's diseases (Lipton & Rosenberg, N. Engl. J. Med. 330, 613-622 (1994)).
Neuropsychiatric disorders are common in cobalamin deficiency and often appear with minimal evidence of hematological abnormality; many of these conditions improve upon cobalamin administration (Lindenbaum et al., N. Engl. J. Med. 318, 1720-1728 (1988)). Some cases of schizophrenia have responded to treatment with vitamin B12 (Regland et al., J. Neural Transm. 98, 143-152 (1994)) or with B12 in combination with other B vitamins (Joshi et al., J. Orthomol. Psychiatry 9, 35-40 (1980)), even in the absence of overt cobalamin deficiency. An association among violent behavior, learning disabilities and low levels of cobalamin in hair samples has been described (Schrauzer et al., Biol. Tr. Element Res. 34, 161-176 (1992)). Since learning disabilities and aggression are frequent concomitants of attention deficit disorder (ADD) (Hallowell & Ratey, Driven to Distraction (New York: Pantheon Books, 1994)), the latter findings suggest a pathogenic role for cobalamin deficiency in ADD, a conclusion supported by reports that vitamin B12 administration is of benefit in treating distractibility and inattention in students (Robin, Semaine hop. Paris 30, 4129-4132 (1954)). Also low serum cobalamin levels have been found among cases of obsessive compulsive disorder (OCD) at a much higher frequency than among controls (Hermesh et al., Acta Psych. Scand. 78, 8-10 (1988)). OCD is believed to result from dysfunction of central serotonergic mechanisms, and abnormal serotonin metabolism has been found in cobalamin deficiency (Botez et al., Ann. Neurol. 12, 479-484 (1982)). Derangements involving serotonin and other neurotransmitters, such as norepinephrine (Deana et al., Int. J. Vit. Nutr. Res. 47, 119-122 (1977)) and GABA (Brennan et al., Brain Res. 219, 186-189 (1981)), may account for many of the mental, psychological and psychiatric disturbances known to accompany cobalamin deficiency. In addition, changes in thyroid function have been reported to be associated with onset of or recovery from depression (Levitt & Joffe, Biol. Psychiatry 33, 52-53 (1993)). Cobalamin deficiency is also known to be linked with depression; recently, the severity of depression in an outpatient population has been positively correlated with serum thyroxine and negatively correlated with serum cobalamin, suggesting an inverse relationship between thyroid hormone and vitamin B12 in the regulation of mood (Levitt & Joffe, op. cit.).
Abnormalities in cobalamin metabolism are often observed in cases of endocrine dysfunction. Serum cobalamin levels are significantly lower in patients with thyrotoxicosis than in controls, suggesting an increased metabolic need for cobalamin in the presence of high levels of thyroid hormone (Alperin et al., Blood 36, 632-641 (1970)). In animal models of thyrotoxicosis, cobalamin supplementation has been shown to counteract the impairment of oxidative phosphorylation and mitochondrial integrity caused by excess thyroid hormone (Kasbekar et al., Biochem. J. 72, 374-383 (1959)). A similar mutual antagonism has been demonstrated between cobalamin and corticosteroids. Thus, vitamin B12 administration counteracts a number of catabolic actions of cortisone in animals (Feng & Meites, Fed. Proc. 14, 47 (1955); Chemnitius, Int. Z. Vitaminforsch. 32, 386-391 (1962)), while experimentally induced cobalamin deficiency results in adrenocortical hypertrophy and elevated serum corticosteroids (Mgongo et al., Reprod. Nutr. Develop. 24, 845-854 (1984)). The latter study also revealed aberrations in gonadal steroid hormone levels consequent to cobalamin deficiency, an effect which may be related to the observed decline in serum B12 among users of estrogen-containing oral contraceptives (Mooij et al., Contraception 44, 277-288 (1991)).
Infertility in both males (Blair et al., Lancet 1, 49-50 (1968)) and females (Menachem et al., Am. J. Hematol. 46, 152 (1994)) has been noted in conjunction with cobalamin deficiency, with restoration of fertility commencing upon treatment with vitamin B12. Moreover, it has long been known that serum cobalamin levels tend to fall during pregnancy (Metz et al., Am. J. Hematol 48, 251-255 (1995)). Recently it has been suggested that a derangement of maternal homocysteine metabolism is responsible for some cases of reproductive disorders such as infertility, recurrent miscarriage and neural tube defects (NTD) (Steegers-Theunissen et al., Fertil. Steril. 60, 1006-1010 (1993)), and that periconceptional supplementation with vitamin B12 may be required for fully effective prophylaxis of NTD (Mills et al., Lancet 345, 149-151 (1995)). Congenital heart defects have been similarly linked with homocysteine embryotoxicity (Rosenquist et al., Proc. Natl. Acad. Sci. USA 93, 15227-15232 ( 1996)). An increased incidence of NTD and other malformations is also known to occur when drugs such as anticonvulsants are administered early in pregnancy; animal studies have demonstrated a role for cobalamin treatment in reducing the incidence of such birth defects (Mann & Gautieri, Lancet 1, 1451-1452 (1973); Elmazar et al., Fund. Appl. Toxicol. 18, 389-394 (1992)).
A novel application for cobalamin therapy is suggested by the finding that vitamin B12 deficiency is associated with increased risk of osteoporosis and bone fractures (Eastell et al., Clin. Sci. 82, 681-685 (1992); Goerss et al., J. Bone Miner. Res. 7, 573-579 (1992)). Marked reversal of bone loss upon treatment with a regimen incorporating cyanocobalamin has been noted (Melton & Kochman, Metabolism 43, 468-469 (1994)). Osteoporosis is also known to be induced by administration of thyroid hormone (Schneider et al., JAMA 271, 1245-1249 (1994)) and corticosteroids (Sambrook & Jones, Br. J. Rheumatol. 34, 8-12 (1995)). As discussed previously, hormone administration or endocrine hyperfunction can deplete cobalamins and, conversely, cobalamin treatment can normalize metabolic imbalances caused by some hormones. In view of these facts, cobalamin therapy is likely to be of use in preventing or reversing hormone-induced osteoporosis as well as the osteoporosis due to cobalamin malabsorption.
Immunodeficiency associated with cobalamin depletion, such as impaired antibody response to pneumococcal vaccine, has been studied in otherwise immunocompetent elderly patients with low serum cobalamin (Fata et al., Ann. Intern. Med. 124, 299-304 (1996)). Previous studies in patients with megaloblastic anemia have shown that immunoglobulin deficiency can resolve upon treatment with vitamin B12 (van Dommelen et al., Acta Med. Scand. 174, 193-200 (1963)). Decreases in suppressor T lymphocyte numbers among pernicious anemia patients have also been found, with one report indicating normalization of CD8+T-cell counts upon administration of hydroxocobalamin (Kubota et al., Am. J. Hematol. 24, 221-223 (1987)). Other impairments of immune function observed in cobalamin deficiency include defective chemiluminescence and bactericidal activity of neutrophils, with microbicidal activity returning to normal after treatment with vitamin B12 (Skacel & Chanarin, Br. J. Haematol. 55, 203-215 (1983)) or hydroxocobalamin (Seger et al., J. Inher. Metab. Dis. 3, 3-9 (1980)). The high incidence of tuberculosis among a vegetarian Indian population in England has been ascribed to defective macrophage killing secondary to dietary cobalamin deficiency, and it has been hypothesized that chronic cobalamin deficiency may particularly predispose individuals to infection by mycobacteria such as those causing tuberculosis and leprosy (Chanarin & Stephenson, J. Clin. Pathol. 41, 759-762 (1988)). The latter hypothesis may be relevant to the recently observed increased prevalence of tuberculosis among vulnerable populations worldwide, especially among those coinfected with HIV (Dolin et al., Bull. World Health Organ. 72, 213-220 (1994)). Thus, there appears to be a specific benefit for cobalamin supplementation in enhancing immunocompetence generally and in the treatment or prophylaxis of mycobacterial infection, of infections due to other microbial pathogens, and of opportunistic infections in AIDS.
Cobalamin deficiency appears to be common both in AIDS patients (Harriman et al., Arch. Intern. Med. 149, 2039-2041 (1989)) and in patients with asymptomatic HIV infection (Rule et al., Am. J. Hematol. 47, 167-171 (1994)); in either case cobalamin malabsorption or depletion has been shown to occur at a very early stage in HIV infection. Other researchers have established that whereas persistent cobalamin deficiency is associated with disease progression, normalization of serum cobalamin levels is associated with increased CD4+T-cell counts and improved AIDS index (a composite measure of disease progression) over time (Baum et al., AIDS 9, 1051-1056 (1995)). These findings suggest that cobalamin deficiency may have a pathogenic role in establishing HIV infection or in progression to AIDS, a supposition supported by the recent discovery that hydroxocobalamin, methylcobalamin and adenosylcobalamin are potent inhibitors of HIV infection in vitro (Weinberg et al., Blood 86, 1281-1287 (1995)). Demyelination, peripheral neuropathy, cognitive and affective changes and dementia are not uncommon consequences of HIV infection, and bear considerable resemblance to the neurological sequelae of chronic vitamin B12 deprivation. A relatively high prevalence of impaired B12 absorption or deficient serum cobalamin levels has been found among HIV-infected patients with neuropathy or myelopathy; the majority of those treated with cyanocobalamin reported a therapeutic response (Kieburtz et al. Arch. Neurol., 48, 312-314 (1991)). The reversal with vitamin B12 of an apparent advanced AIDS dementia complex has also been reported (Herzlich & Schiano, J. Intern. Med. 233, 495-497 (1993)), while measures of cognitive functioning have been directly correlated with serum cobalamin levels in HIV-infected patients (Shor-Posner et al., Arch. Neurol. 52, 195-198 (1995)).
Various cobalamins have shown efficacy in inhibiting tumor cell growth in culture, and in treating neoplasms and premalignant lesions in individuals. Pernicious anemia has been associated with an elevated risk of cancer, including melanoma, multiple myeloma, myeloid and other leukemias, and oral, pharyngeal and gastric cancers; the declining risk of leukemia and gastric carcinoma from the time of diagnosis of pernicious anemia is thought to result from therapy with vitamin B12 (Brinton et al., Br. J. Cancer 59, 810-813 (1989)). Deficiency of cobalamin and/or folate is believed to play a procarcinogenic role in general by impairing methionine synthesis, thereby inducing DNA hypomethylation (Herbert, in Essential Nutrients in Carcinogenesis (New York: Plenum Press, 1986), 293-311). Thus, dietary deficiency of methyl donors such as methionine and folate has been linked with increased risk of colorectal adenoma (Giovannucci et al., J. Natl. Cancer Inst. 85, 875-884 (1993)), and low serum cobalamin and folate levels have been correlated with a high incidence of esophageal carcinoma (EC) (Ran et al., Blood C, Suppl. 82, 532a (1993)). The latter authors have also shown that dietary supplementation with cobalamin and folate can correct esophageal dysplasia, the immediate precursor of EC, in cobalamin- and folate-deficient individuals. Similarly, smoking has been reported to induce a localized cobalamin deficiency in mucosal tissues (Piyathilake et al., FASEB 7, 713 (1993)), whereas treatment with vitamin B12 and folate has been shown to improve bronchial squamous metaplasia, a precursor of lung cancer, among smokers (Heimburger et al., JAMA 259, 1525-1530 (1988)). Moreover, when coadministered with folate, cobalamin has been reported to potentiate fluoropyrimidine antitumor activity (Tisman et al., Clin. Res. 33, 459A (1985)). There are also reports that cases of neuroblastoma (Bodian, Arch. Dis. Child. 38, 606-619 (1963)) and retinoblastoma (Horne, Am. J. Ophthalmol. 61, 910-911 (1966)) have responded to therapy with vitamin B12. In addition, studies conducted in vitro with the coenzyme forms of vitamin B12 have shown a cytotoxic effect of adenosylcobalamin on fast-growing malignant cell lines, with a lesser cytotoxicity induced by methylcobalamin (Tsao et al., Pathobiology 58, 292-296 (1990)); the cytotoxicity of adenosylcobalamin may be due to its inhibition of tRNA methylase activity, which is known to be elevated in tumor tissues (Tarasyavichene et al., Biokhimiya 41, 1614-1618 (1976)). These latter results suggest the existence of at least two distinct and independent mechanisms for the procarcinogenic effects of cobalamin deficiency.
Many of the aforementioned conditions associated with cobalamnin deficiency--e.g., AIDS, diabetes, atherosclerosis, apoptotic conditions, autoimmune and chronic inflammatory diseases, neurodegenerative conditions--are also associated with oxidative stress and/or antioxidant depletion. Cobalamin deficiency has itself been linked with alterations in redox status of endogenous antioxidants such as glutathione (GSH), ascorbic acid and reduced nicotinamide adenine dinucleotide (NADH). For example, elevated levels of oxidized GSH have been found in erythrocytes from untreated pernicious anemia patients, with improvement in redox status following upon treatment with vitamin B12 (Jocelyn, Biochem. J. 77, 363-368 (1960)). Similarly, an abnormally rapid oxidation of plasma ascorbic acid to dehydroascorbic acid has been noted in some cases of pernicious anemia, with the abnormality disappearing after cobalamin supplementation (Will et al., J. Lab. Clin. Med. 42, 967 (1953); Mueller and Will, Am. J. Clin. Nutr. 3, 30-44 (1955)). Other aberrations in ascorbic acid metabolism have also been observed in cobalamin deficiency (Cox et al., Clin. Sci. 17, 681-692 (1958)). Decreases in GSH (Register, J. Biol. Chem. 206, 705-709 (1954)), GSH reductase activity (Biswas & Johnson, Arch. Biochem. Biophys. 104, 375-380 (1964)) and in the ratio NADH/NAD (Chang et al., Fed. Proc. 16, 163-164 (1957)) have been found in livers of cobalamin-deficient animals. Thus, vitamin B12 appears to play a key role in maintaining antioxidants in their reduced state and/or in facilitating their proper metabolism. This conclusion may be generally relevant to the treatment of oxidative stress, inasmuch as oxidative stress can induce a functional cobalamin deficiency by oxidatively degrading the cobalamin molecule (e.g., via hydroxyl radical attack on the corrin ring) to yield toxic or inactive cobalamin analogues (Nazhat et al., J. Inorgan. Biochem. 36, 75-81 (1989)).
In particular, cobalamin supplementation alone or in combination with GSH may be beneficial in the treatment of oxidative stress associated with methylmercury intoxication, as has been demonstrated for various B complex vitamins and GSH (Sood et al., Cell. Mol. Biol. 39, 213-219 (1993)). Cobalamin may also be useful in the treatment of conditions characterized by both oxidative stress and glutamatergic excitotoxicity, such as diabetic retinopathy, macular degeneration and Batten's disease, among others (Agostinho et al., FASEB J. 11, 154-165 (1997)); a protective effect of cobalamin against glutamatergic excitotoxicity has been previously discussed (Yamamoto et al., op. cit.). Other conditions of impaired antioxidant homeostasis where cobalamin supplementation may be of use include cataract, heart disease (Harding et al., Biochem. Soc. Trans. 24, 881-883 (1996)) and also prolonged physical exercise (Reid et al., J. Cliln. Invest. 94, 2468-2474 (1994); Leeuwenburgh & Ji, Arch. Biochem. Biophys. 316, 941-949 (1995)). With or without the accompanying administration of antioxidants, cobalamin may be useful in inhibiting the muscle fatigue induced by prolonged exercise (Reid et al., op. cit.); indeed, studies conducted among ultraendurance athletes have revealed metabolic abnormalities suggesting an increased need for vitamin B12 in such athletes (Singh et al., Med. Sci. Sports Exerc. 25, 328-334 (1993)).
In addition to the various uses of cobalamin in treating conditions associated with cobalamin deficiency, cobalamins are effective in a number of applications regardless of whether any deficiency exists. Many of these applications stem from the avidity with which cobalamins scavenge, oxidize or otherwise interact with small bioactive molecules such as cyanide, nitric oxide, superoxide, carbon monoxide, sulfite and also with halogenated hydrocarbons. For example, hydroxocobalamin has long been known as an effective antidote for cyanide poisoning (Mushett et al., Proc. Soc. Exp. Biol. Med. 81, 234-237 (1952); Zerbe & Wagner, Crit. Care Med. 21, 465-467 (1993)). Recently it has been determined that cobalamins also interact with nitric oxide (Weinberg et al., Blood 84, 118a (1994)), and that hydroxocobalamin in particular attenuates the nitric oxide-dependent hypotension and mortality induced by septic shock (Greenberg et al., J. Pharmacol. Exp. Ther. 273, 257-265 (1995)). The latter authors suggest the use of hydroxocobalamin for treatment of sepsis, endotoxemia, systemic inflammatory response syndrome and other disorders associated with excess production of nitric oxide, as well as for adjunctive therapy when coadmninistered with inhaled nitric oxide or with nitric oxide donors. In addition to autoimmune disorders, chronic inflammatory disease, neurodegenerative disease and HIV infection, other conditions of nitric oxide toxicity which may be mitigated by cobalamins include migraine (Olesen et al., NeuroReport 4, 1027-1030 (1993)), stroke (Nowicki et al., Eur. J. Pharmacol. 204, 339-340 (1991)), viral pneumonia (Akaike et al., Proc. Natl. Acad. Sci. USA 93, 2448-2453 (1996)) and viral and bacterial neurological disease (Zheng et al., J. Virol. 67, 5786-5791 (1993); Koedel et al., Ann. Neurol. 37, 313-323 (1995)).
Similarly, a cobalamin complex with superoxide anion has been described (Bayston et al., J. Am. Chem. Soc. 91, 2775-2779 (1969)). Superoxide is a reactive oxygen species often coreleased with nitric oxide and implicated with it in the pathogenesis of AIDS and of various autoimmune, chronic inflammatory, ischemic and neurodegenerative diseases. With or without an accompanying release of nitric oxide, excessive production of superoxide has been linked with numerous pathologies, including infection by viral, bacterial, parasitic and fingal pathogens (Fuchs et al., Med. Hypotheses 36, 60-64 (1991)), induction of muscle wasting in cachexia (Buck & Chojkier, EMBO J. 15, 1753-1765 (1996)), photodamage to skin (Darr & Fridovich, J. Invest. Dermatol. 102, 671-675 (1994)), and the generation of clastogenic factors in a variety of illnesses. Clastogenic factors are low molecular weight chromosome-damaging agents which cause chromosome aberrations, sister chromatid exchanges, DNA strand breakage and gene mutation; their production is induced by superoxide and they in turn promote the formation of additional superoxide (Fuchs et al., Free Radic. Biol. Med. 19, 843-848 (1995)). Clastogenic factors have been implicated in the pathogenesis of gene mutations and/or malignancies associated with exposure to ionizing radiation, viruses, tumor-promoting chemicals, asbestos, herbicides such as paraquat, and with hereditary chromosome breakage syndromes such as ataxia telangiectasia, Bloom's syndrome and Fanconi's anemia (Emerit, Free Radic. Biol. Med. 16, 99-109 (1994)). All of the conditions cited represent targets for mitigation of superoxide toxicity by cobalamin.
Carbon monoxide, sulfites and various halogenated hydrocarbons also interact with cobalamin. Cobalamins have been reported to catalyze the oxidation of carbon monoxide to carbon dioxide (Bayston & Winfield, J. Catalysis 9, 217-224 (1967); Thauer et al., Eur. J. Biochem. 45, 343-349 (1974)) with potentially significant implications for the treatment of conditions associated with carbon monoxide production or exposure, such as smoking in adults and fetal growth retardation, sudden infant death syndrome (Hutter & Blair, Med. Hypotheses 46, 1-4 (1996)) and other pediatric conditions (Stevenson et al., J. Pediatrics 94, 956-958 (1979)). In addition, the neurotoxicity of carbon monoxide poisoning has been linked with excess nitric oxide production (Ischiropoulos et al., J. Clin. Invest. 97, 2260-2267 (1996)), which can be attenuated by hydroxocobalamin administration as previously noted. Cyanocobalamin is also known to be effective in the treatment of asthma in general (Crocket, Acta Allergol. 11, 261-268 (1957); Wright, Int. Clin. Nutr. Rev. 9, 185-188 (1989)) and in the suppression of allergic reactions to sulfites in cases of sulfite-sensitive asthma (SSA) in particular (Anibarro et al., J. Allergy Clin. Irnmunol. 90, 103-109 (1992)). The protective effect of vitamin B12 in non-SSA asthma may be due to the scavenging of nitric oxide by cobalamin, since excess production of nitric oxide is known to be involved in asthmatic inflammation (Lundberg et al., Nature Med. 3, 30-31 (1997)). In contrast, the protective effect of cyanocobalamin in SSA is presumed to be a consequence of the extracellular nonenzymatic oxidation of sulfite catalyzed by cobalamins (Jacobsen et al., J. Allergy Clin. Immunol. 73, 135 (1984)). Finally, vitamin B12 has been shown to mediate the dehalogenation of various halogenated pesticides (Schrauzer & Katz, Bioinorg. Chem. 9, 123-143 (1978)), environmental toxins (Assaf-Anid et al., Appl. Env. Microbiol. 58, 1057-1060 (1992)) and solvents (Krone et al., Biochemistry 30, 2713-2719 (1991)), a result which may account for the protection afforded by vitamin B12 in carbon tetrachloride-induced hepatic injury (Kasbekar et al., Biochem. J. 72, 384-389 (1959)).
Other applications for cobalamins include treatment of dermatitis (Simon, J. Allergy 22, 183-185 (1951)); antagonism of histamine (Ata, in Vitamin B12 und Intrinsic Factor (Stuttgart: Ferdinand Enke Verlag, 1957), 544-553); treatment of oversedation due to intoxication with sedatives and/or alcohol (Newbold, Med. Hypotheses 30, 1-3 (1989)); treatment of anorexia nervosa (Korkina et al., Zhur. Nevropat. Psikhiat. 89, 82-87 (1989)); relief of fatigue (Ellis & Nasser, Br. J. Nutr. 30, 277-283 (1973)); enhancement of choline and acetylcholine biosynthesis (Sasaki et al., Pharmacol. Biochem. Behav. 43, 635-639 (1992)); treatment of sleep disturbances and "jet lag" by re-entrainment of circadian rhythms (Honma et al., Experientia 48, 716-720 (1992)); treatment of viral conditions such as hepatitis (Kelemen et al., Int. Z. Vitaminforsch. 31, 307-316 (1961)), poliomyelitis (Leroy & Robin, Semaine hop. Paris 31, 1097-1098 (1955)), and herpetic lesions (King, N.Z. Med. J. 105, 135 (1992)); potentiation of immunomodulation when coadministered with interferon (Medenica et al., Blood 86, Suppl. 1, 850a (1995)); pain relief (Surtees & Hughes, Lancet 1, 439-441 (1954); Leuschner, Arzneim.-Forsch./Drug. Res. 42, 114-115 (1992)); treatment of osteoarthritis (Flynn et al., J. Am. Coll. Nutr. 13, 351-356 (1994)); promotion of epithelial cell growth (Ansell, Lancet 2, 994 (1962)), of wound healing (Findlay, Proc. Soc. Exp. Biol. Med. 82, 492-495 (1953)) and of recovery of cardiac muscle in myocardial infarction (Nikolaeva et al., Circ. Res. 35, Suppl. III, 202-213 (1974)); detoxification of poisoning caused by heavy metals such as cadmium (Couce et al. J. Inorg. Biochem. 41, 1-6 (1991)), lead (Kleinsorge et al., Zschr. inn. Med. 9, 903-906 (1954)) and mercury (e.g., methylmercury, Sood et al., Cell. Mol. Biol. 39, 213-219 (1993)) and non-metals such as selenium (Chen & Whanger, Toxicol. Appl. Pharmacol. 18, 65-72 (1993)); antagonism of convulsions and mortality caused by various agents and medications (Ata, in Vitamin B12 und Intrinsic Factor (Stuttgart: Ferdinand Enke Verlag, 1957), 544-553) and treatment of febrile convulsions (Osifo et al., J. Neurol. Sci. 68, 185-190 (1985)).
Corrinoids are cobalamin analogs which are used in lieu of cobalamin by certain microorganisms. Generally, corrinoids differ from cobalamins only in an alteration of the dimethylbenzimidazole moiety found in all true cobalamins. Corrinoids are believed to be inactive in eukaryotic cells, and there is evidence that corrinoids may actually antagonize some functions of vitamin B12 or otherwise interfere with cobalamin uptake or metabolism in humans and animals. Both cobalamins and corrinoids may nevertheless be useful in eukaryotes or, e.g., as growth factors, prebiotics or essential nutrients for microorganisms of value, such as those employed in fermentation reactions or in environmental detoxification, for example.